Cryopreservation of in Vitro Grown Shoot Tips of Sweet Potato (Ipomoea batatas L.) by the Encapsulation-Vitrification Method

Abstract

-Sweet potato (Ipomoea batatas L.) shoot tips grown in vitro were successfully cryopreserved by encapsulationvitrification. Encapsulated explants are very easily manipulated, due to the relatively large size of the alginate beads, and a large number of samples can be treated simultaneously. In this study, the effects of sucrose preculture, cryoprotectant preculture, and post-warm recovery media on regrowth, following liquid nitrogen (LN) exposure, were investigated to establish an efficient encapsulation-vitrification protocol for sweet potato. Shoot tips of plants grown in vitro were precultured in 0.3 M sucrose for 2 d before encapsulation. Encapsulated shoot tips were pre-incubated in liquid MS (Murashige and Skoog) medium containing 0.5 M sucrose for 16 h, before preculturing in sucrose-enriched medium (0.7 M sucrose) for 8 h. Shoot tips were osmoprotected with 35% plant vitrification solution 3 (PVS3) for 3 h, before being dehydrated with PVS3 for 2 h at 25°C. The encapsulated and dehydrated shoot tips were transferred to 2 mL cryotubes, suspended in 0.5 mL PVS3, and plunged directly into liquid N. High levels of shoot formation were obtained for the cv. Yeulmi (65.7%) and Yeonwhangmi (80.3%). The regrowth rates of cryopreserved samples in Yeulmi (78.9%) and Yeonwhangmi (91.3%), following culture on ammonium-free MS medium for 5 d, were much higher than those cultured on standard MS medium (65.7% and 80.3%, respectively). This encapsulation-vitrification is a promising method for the long-term preservation of sweet potato.

Overcoming Challenges for Shoot Tip Cryopreservation of Root and Tuber Crops

“…Although the En-vi has been successfully applied to many plant species, this method is less studied compared to other cryopreservation methods for RTCs. Even though this technique has been used with a limited number of RTCs, it possesses great potential both in terms of efficiency and practicality [52,53,138].…”

Critical Role of Regrowth Conditions in Post-Cryopreservation of In Vitro Plant Germplasm

“…Ammonium salts are a standard component of in vitro culture medium, but they may be harmful to plant materials experiencing severe stress [91] and can be removed or reduced in quantity for better post-LN regrowth. Even low concentrations of ammonium nitrate may inhibit the normal recovery of shoot tips and undifferentiated cell cultures Table 1 [92][93][94][95]. Removing or substituting ammonium in the regrowth medium improved the regeneration of the cryopreserved shoot tips of Holostemma annulare, Ipomoea batatas, Chrysanthemum morifolium var.…”

“…'Frost Eureca limon' and Aster altaicus var. uchiyamae [93][94][95][96][97][98]. Regeneration improvement compared to the ammonium-containing medium varied from 17 to over 60% depending on species/plant material; 30-40% improvement was the most common (Table 1).…”

Biobanking of vegetable genetic resources by in vitro conservation and cryopreservation

“…Since 2000, four studies have optimized two different procedures for the cryopreservation of shoot tips (around 1 mm in size) taken from in vitro shoot cultures of sweet potato: droplet vitrification and encapsulation-vitrification, using either PVS2 or PVS3 as vitrification solutions. It should be noted that, in these studies, the rewarming procedure applied to the explants recovered from LN was quite different, as well as the percentages of shoot tip regrowth, ranging from a minimum of 19% (Park and Kim 2015) to a maximum of 91% (Yi et al 2016). However, this variability in the outcomes of the procedures did not prevent the introduction of the species into conservation in some cryobanks (Table 3).…”